Global analysis of lysine 2-hydroxyisobutyrylation in wheat root

Abstract

Lysine 2-hydroxyisobutyrylation (Khib) is a novel naturally occurring post-translational modification. The system Khib identification at proteomics level has been performed in various species and tissues to characterize the role of Khib in biological activities. However, the study of Khib in plant species is relatively less. In the present study, the first plant root tissues lysine 2-hydroxyisobutyrylome analysis was performed in wheat with antibody immunoprecipitation affinity, high resolution mass spectrometry-based proteomics and bioinformatics analysis. In total, 6328 Khib sites in 2186 proteins were repeatedly identified in three replicates. These Khib proteins showed a wide subcellular location distribution. Function and pathways characterization of these Khib proteins indicated that many cellular functions and metabolism pathways were potentially affected by this modification. Protein and amino acid metabolism related process may be regulated by Khib, especially ribosome activities and proteins biosynthesis process. Carbohydrate metabolism and energy production related processes including glycolysis/gluconeogenesis, TCA cycle and oxidative phosphorylation pathways were also affected by Khib modification. Besides, root sulfur assimilation and transformation related enzymes exhibited Khib modification. Our work illustrated the potential regulation role of Khib in wheat root physiology and biology, which could be used as a useful reference for Khib study in plant root.

Figure 1

Systematic analysis of Khib in wheat roots. (A) Workflow for global Khib screening in wheat roots. (B) Mass error distribution of all the Khib peptides. (C) Peptide length distribution of all the Khib peptides. (D) Venn diagram of the identified Khib sites. (E) Venn diagram of the identified Khib proteins.

Figure 2

Functional classification and subcellular location analyses of the Khib proteins. (A) Function classification based on GO annotation. (B) Subcellular location from WoLF PSORT.

Figure 3

Enrichment analyses of the Khib proteins. (A) GO annotation based enrichment analysis. (B) KEGG pathway based enrichment analysis.

Figure 4

Representative significantly enriched protein metabolism related KEGG pathways. (A) Ribosome. (B) Proteasome. The Khib proteins are labeled in red.

Figure 5

Representative significantly enriched carbohydrate metabolism and energy production related KEGG pathways. (A) Glycolysis/gluconeogenesis. (B) TCA cycle. (C) Oxidative phosphorylation. The Khib proteins are labeled in red.

Figure 6

Khib regulated sulfur assimilation and metabolism. The Khib modified enzymes are indicated in green. APS: adenosine 5′-phosphosulfate; PAPS: adenosine 3′-phosphate 5′-phosphosulfate; PAPSS: Sulfate adenylyltransferase; CYSH: phosphoadenosine phosphosulfate reductase; CYSQ: 3′(2′),5′-bisphosphate nucleotidase, SELENBP1:Selenium-binding protein 1, methanethiol oxidase; TST: thiosulfate/3-mercaptopyruvate sulfurtransferase; TSR: thiosulfate-dithioerythritol sulfurtransferase; PSRA: thiosulfate reductase/polysulfide reductase chain A.